1. Field of the Invention
The present invention relates to assays and methods of use, and more particularly, to detection of polarized angular scattering from plasmonic nanostructures for determining concentrations of receptor-ligand binding.
2. Background of the Related Art
Typically, in cellular imaging today, fluorophores or even quantum dots, are used, which either contain some function groups to bind to expressed cellular surface features (receptors) or can even be transfected within the cells. This enables the cells to be readily imaged. However, one particular problem with using fluorophores is there inherent photo instability, where most fluorophores typically photo degrade after about 103 excitation/emission event cycles.
In spectroscopy today, the techniques are typically limited by the wavelength of light for imaging, structures and features less than 1 micron in size being most difficult to see. However, contrary to this, FRET (Fluorescence Resonance Energy Transfer) has earned a well deserved reputation for being able to indirectly image features that are within the Forster Transfer distances of fluorophores, that being, 5 nm. Hence, FRET is widely used to study macromolecular dynamics. However, there is a diffraction limited gap in imaging today because current technology cannot image structures and features in the 10 to 1000 nm size.
Over the last several years, the use of both gold and silver nanoparticles in biological assays has dramatically increased. Nanostructures are far superior to fluorophores in that they don't photodegrade and have “Plasmon Scattering Powers” far greater than the emission of fluorophores. This has been afforded by their very high molar absorption coefficients. In addition to their high absorption cross-sections, nanoparticles of gold and silver are also very efficient scatterers of light. Indeed a noble metal colloid's extinction spectrum is composed of both an absorption and scattering component, which is contrary to how we think of a typical fluorophores extinction spectrum. Subsequently, light scattering by gold and silver nanoparticles can be detected at concentrations as low as 10−16 M. For example, a 20 nm gold colloid can scatter light at 532 nm, the equivalent intensity as 105 fluorescing fluorescein molecules. In addition, it is well known that the light dependent scattering properties of nanoparticles depend on their size, shape, composition and the refractive index of the suspending medium. However, one property that has been ill explored for biosensing applications is the polarization and intensity of plasmon scatter.
Thus, it would be advantageous to provide a method for using polarized scatter from plasmonic nanostructures for bioassays.